Stewart S, Yi S, Kassabian G, Mayo M, Sank A, Shuler C
Department of Craniofacial Molecular Biology, University of Southern California, Los Angeles, CA 90033, USA.
Anat Embryol (Berl). 2000 Jun;201(6):483-90. doi: 10.1007/s004290050335.
Syndactyly, a failure of the digits to separate into individual units, affects about 8 to 9 per 1000 newborns and results from an aberration of the normal development of the interdigital tissues. Limb digit separation is the result of programmed cell death (apoptosis). Lysosomes play a role in the process of cell self-destruction. Our experiment was designed to test the hypothesis that the intensity of interdigital lysosomes increases during the separation of digits in vivo and in vitro. The primary mouse monoclonal antibody, 1D4B, detects the presence of lysosomes by identifying the LAMP-1 glycoprotein on the lysosome cell membrane. In our experiment this antibody immunodetected interdigital lysosome proteins in serial sections of limbs from Swiss-Webster mouse embryos, gestational ages E12.5 through E15, key developmental stages for digit separation. Digit separation was associated with an increase in intensity of lysosomal protein staining. In E12.5 limbs, the presence of lysosomes was enriched in the distal aspect of the interdigital tissue. However, the number of lysosomes markedly increased in the E13 and E14 limbs, including the entire length and width of the interdigital tissue in the E14 limbs. This lysosomal protein presence in E14 limbs was significant compared to E12.5, E13, and E15 limbs. By day 12.5, the mouse embryo limb is committed to digit separation. Addition of retinoic acid to the culture medium of limbs earlier in development, such as E12, results in induction of the process of digit separation. Cultured E12 limbs that did not receive an addition of retinoic acid, did not show digit separation. We conclude that in the limb development process, the enrichment in interdigit LAMP-1 proteins, may indicate a relationship between lysosomes, apoptosis, and digit separation. We also conclude that retinoic acid has an important role in digit separation in vivo, as shown in limb development, and demonstrated through the addition of retinoic acid to media of cultured tissues.
并指是一种手指未能分离成独立个体的病症，每1000名新生儿中约有8至9人受其影响，它是由指间组织正常发育异常所致。肢体手指分离是程序性细胞死亡（凋亡）的结果。溶酶体在细胞自我毁灭过程中发挥作用。我们的实验旨在验证这样一个假设：在体内和体外手指分离过程中，指间溶酶体的强度会增加。小鼠单克隆抗体1D4B通过识别溶酶体细胞膜上的LAMP - 1糖蛋白来检测溶酶体的存在。在我们的实验中，这种抗体对瑞士 Webster 小鼠胚胎（妊娠年龄从E12.5到E15，这是手指分离的关键发育阶段）肢体的连续切片中的指间溶酶体蛋白进行了免疫检测。手指分离与溶酶体蛋白染色强度增加有关。在E12.5的肢体中，溶酶体在指间组织的远端富集。然而，在E13和E14的肢体中，溶酶体数量显著增加，在E14的肢体中包括指间组织的整个长度和宽度。与E12.5、E13和E15的肢体相比，E14肢体中这种溶酶体蛋白的存在是显著的。到第12.5天，小鼠胚胎肢体开始进行手指分离。在发育早期，如E12时，向肢体培养基中添加视黄酸会导致手指分离过程的诱导。未添加视黄酸的培养E12肢体未出现手指分离。我们得出结论，在肢体发育过程中，指间LAMP - 1蛋白的富集可能表明溶酶体、凋亡和手指分离之间存在关联。我们还得出结论，视黄酸在体内手指分离中具有重要作用，如在肢体发育中所示，并通过向培养组织的培养基中添加视黄酸得到证明。
